Acanthamoeba Infection Research Articles

Amoebicidal and cysticidal in vitro activity of cationic dendritic molecules against Acanthamoeba polyphaga and Acanthamoeba griffini.

Acanthamoeba species are responsible for serious human infections, including Acanthamoeba keratitis (AK) and granulomatous amoebic encephalitis (GAE). These pathogens have a simple life cycle consisting of an infective trophozoite stage and a resistant cyst stage, with cysts posing significant treatment challenges due to their resilience against harsh conditions and chemical agents. Current treatments for AK often involve combining diamines, such as propamidine, and biguanides, such as chlorhexidine (CLX), which exhibit limited efficacy and significant toxicity. Thus, the effect of new therapeutic molecules, such as multifunctional systems (e.g., carbosilane dendritic molecules), should be studied as potential alternatives due to their biocidal properties and lower toxicity. This study evaluates various dendritic compounds against trophozoites and cysts of two Acanthamoeba clinical isolates, both alone and in combination with CLX, and assesses their cytotoxicity on HeLa cells. The results indicated that certain dendritic compounds, especially BDSQ024, were effective against both trophozoites and cysts. Additionally, combinations of dendritic molecules and CLX showed enhanced efficacy in eliminating trophozoites and cysts, suggesting potential for synergistic treatments. The study underscores the promise of dendritic molecules in developing more effective and less toxic therapies for Acanthamoeba infections.

Drug modifications: graphene oxide-chitosan loading enhanced anti-amoebic effects of pentamidine and doxycycline.

Acanthamoeba castellanii is the causative pathogen of a severe eye infection, known as Acanthamoeba keratitis and a life-threatening brain infection, named granulomatous amoebic encephalitis. Current treatments are problematic and costly and exhibit limited efficacy against Acanthamoeba parasite, especially the cyst stage. In parallel to drug discovery and drug repurposing efforts, drug modification isalsoan important approach to tackle infections, especially against neglected parasites such as free-living amoebae: Acanthamoeba. In this study, we determined whether modifying pentamidine and doxycycline through chitosan-functionalized graphene oxide loading enhances their anti-amoebic effects. Various concentrations of doxycycline, pentamidine, graphene oxide, chitosan-functionalized graphene oxide, and chitosan-functionalized graphene oxide loaded with doxycycline and pentamidine were investigated for amoebicidal effects against pathogenic A. castellanii belonging to the T4 genotype. Lactate dehydrogenase assays were performed to determine toxic effects of these various drugs and nanoconjugates against human cells. The findings revealed that chitosan-functionalized graphene oxide loaded with doxycycline demonstrated potent amoebicidal effects. Nanomaterials significantly (p < 0.05) inhibited excystation and encystation of A. castellanii without exhibiting toxic effects against human cells in a concentration-dependent manner, as compared with other formulations. These results indicate that drug modifications coupled with nanotechnology may be a viable avenue in the rationale development of effective therapies against Acanthamoeba infections.

Systemic Acanthamoeba T17 infection in a free-ranging two-toed sloth: case report and literature review of infections by free-living amebas in mammals.

A free-ranging, adult female two-toed sloth (Choloepus hoffmanni) was brought to a wildlife rescue center in Costa Rica with ocular and auricular myiasis and numerous skin lesions. After one month of unsuccessful systemic and topical antimicrobial treatment, the patient died. A postmortem examination was performed, and tissues were examined histologically, confirming disseminated amebic infection with intralesional trophozoites and cysts in the lungs, liver, eye, heart, spleen, and stomach. Immunohistochemistry identified the ameba as Acanthamoeba sp. A multiplex real-time PCR assay, 18S ribosomal DNA PCR, and sequencing performed on formalin-fixed, paraffin-embedded lung tissue confirmed the Acanthamoeba T17 genotype. The Acanthamoeba genus is in the group of free-living amebas that cause infection in humans and animals, and it is ubiquitous in the environment. Acanthamoeba T17 has been isolated from water and soil, but to our knowledge, this genotype has not been implicated in infections of animals previously and has not been reported from Costa Rica. Systemic Acanthamoeba infection has not been described in sloths previously. We provide a comprehensive literature review describing infections by free-living amebas of the genus Acanthamoeba spp., Balamuthia spp., and Naegleria spp. in domestic, zoo, and wild mammals.

Identification of new targets for the diagnosis of cysts (four) and trophozoites (one) of the eye pathogen Acanthamoeba.

Free-living amoeba in the soil or water cause Acanthamoeba keratitis, which is diagnosed by identification of cysts by microscopy of the eye or of corneal scrapings, using calcofluor-white that unfortunately cross-reacts with fungi and plants. Alternatively, Acanthamoeba infections are diagnosed by identification of trophozoites in cultures of scrapings. Here we showed that rabbit antibodies to four abundant cyst wall proteins (Jonah, Luke, Leo, and laccase) each efficiently detect calcofluor-white-labeled cysts of 10 of 11 Acanthamoeba isolates obtained from the ATCC. Further, laccase released into the medium by encysting Acanthamoebae was detected by an enzyme-linked immunoassay. We also showed that rabbit antibodies to the mannose-binding domain of the Acanthamoeba mannose-binding protein, which mediates adherence of trophozoites to keratinocytes, efficiently identifies trophozoites of all 11 ATCC isolates. In summary, four wall proteins and the ManBD appear to be excellent targets for diagnosis of Acanthamoeba cysts and trophozoites, respectively.

Acanthamoeba castellanii trophozoites need oxygen for normal functioning and lipids are their preferred substrate, offering new possibilities for treatment

Acanthamoebae, pathogenic free-living amoebae, can cause Granulomatous Amoebic Encephalitis (GAE) and keratitis, and for both types of infection, no adequate treatment options are available. As the metabolism of pathogens is an attractive treatment target, we set out to examine the energy metabolism of Acanthamoeba castellanii and studied the aerobic and anaerobic capacities of the trophozoites. Under anaerobic conditions, or in the presence of inhibitors of the electron-transport chain, A. castellanii trophozoites became rounded, moved sluggishly and stopped multiplying. This demonstrates that oxygen and the respiratory chain are essential for movement and replication. Furthermore, the simultaneous activities of both terminal oxidases, cytochrome c oxidase and the plant-like alternative oxidase, are essential for normal functioning and replication. The inhibition of normal function caused by the inactivity of the respiratory chain was reversible. Once respiration was made possible again, the rounded, rather inactive amoebae formed acanthopodia within 4 h and resumed moving, feeding and multiplying. Experiments with radiolabelled nutrients revealed a preference for lipids over glucose and amino acids as food. Subsequent experiments showed that adding lipids to a standard culture medium of trophozoites strongly increased the growth rate. Acanthamoeba castellanii trophozoites have a strictly aerobic energy metabolism and β-oxidation of fatty acids, the Krebs cycle, and an aerobic electron-transport chain coupled to the ATP synthase, producing most of the used ATP. The preference for lipids can be exploited, as we show that three known inhibitors of lipid oxidation strongly inhibited the growth of A. castellanii. In particular, thioridazine and perhexiline showed potent effects in low micromolar concentrations. Therefore, this study revealed a new drug target with possibly new options to treat Acanthamoeba infections.

A chorismate mutase-targeted, core-shell nanoassembly-activated SERS immunoassay platform for rapid non-invasive detection of Acanthamoeba infection

Contact lens care and early diagnosis of Acanthamoeba keratitis (AK) are very important to prevent progression to blindness due to AK, which develops when Acanthamoeba attaches to contact lens-damaged corneas. Therefore, we propose a novel, non-invasive, immuno-surface-enhanced Raman scattering (SERS) sensing platform for rapid and accurate detection of Acanthamoeba infection in the tears and contact lens solutions of humans. This optic analysis method was based on the proven biological performance of chorismate mutase (CM)-specific monoclonal and polyclonal antibodies on trophozoite and cyst forms of Acanthamoeba castellanii, and its conditioned media. SERS-based, ultra-low concentration detection was achieved by the anisotropic fanblade-shaped core-shell nanoassembly (Ag@AuFNP) embedded with 4-fluorobenzenethiol Raman reporter. The immuno-SERS platform combining Ag@AuFNP and CM-specific antibody complexes was evaluated in vitro and in vivo. The non-invasive SERS-activated biosensing platform indicates strong feasibility for AK detection in human tears and contact lens solutions.

Staurosporine as a Potential Treatment for Acanthamoeba Keratitis Using Mouse Cornea as an Ex Vivo Model.

Acanthamoeba is a ubiquitous genus of amoebae that can trigger a severe and progressive ocular disease known as Acanthamoeba Keratitis (AK). Furthermore, current treatment protocols are based on the combination of different compounds that are not fully effective. Therefore, an urgent need to find new compounds to treat Acanthamoeba infections is clear. In the present study, we evaluated staurosporine as a potential treatment for Acanthamoeba keratitis using mouse cornea as an ex vivo model, and a comparative proteomic analysis was conducted to elucidate a mechanism of action. The obtained results indicate that staurosporine altered the conformation of actin and tubulin in treated trophozoites of A. castellanii. In addition, proteomic analysis of treated trophozoites revealed that this molecule induced overexpression and a downregulation of proteins related to key functions for Acanthamoeba infection pathways. Additionally, the ex vivo assay used validated this model for the study of the pathogenesis and therapies of AK. Finally, staurosporine eliminated the entire amoebic population and prevented the adhesion and infection of amoebae to the epithelium of treated mouse corneas.

Metal Oxide Nanoparticles Exhibit Anti-Acanthamoeba castellanii Properties by Inducing Necrotic Cell Death.

The treatment of amoebic infections is often problematic, largely due to delayed diagnosis, amoebae transformation into resistant cyst form, and lack of availability of effective chemotherapeutic agents. Herein, we determined anti-Acanthamoeba castellanii properties ofthree metal oxide nanoparticles (TiO2, ZrO2, and Al2O3). Amoebicidal assays were performed to determine whether metal oxide nanoparticles inhibit amoebae viability. Encystation assays were performed to test whether metal oxide nanoparticles inhibit cyst formation. By measuring lactate dehydrogenase release, cytotoxicity assays were performed to determine human cell damage. Hoechst 33342/PI staining was performed to determine programmed cell death (apoptosis) and necrosis in A. castellanii. TiO2-NPs significantly inhibited amoebae viability as observed through amoebicidal assays, as well as inhibited their phenotypic transformation as evident using encystation assays, and showed limited human cell damage as observed by measuring lactate dehydrogenase assays. Furthermore, TiO2-NPs altered parasite membranes and resulted in necrotic cell death as determined using double staining cell death assays with Hoechst33342/Propidium iodide (PI) observed through chromatin condensation. These findings suggest that TiO2-NPs offers a potential viable avenue in the rationaledevelopment oftherapeutic interventions against Acanthamoeba infections.

Evaluating the thioredoxin reductase selenoprotein as potential drug target for treatment of Acanthamoeba infections

The genus Acanthamoeba comprises facultative pathogens, causing Acanthamoeba keratitis (AK) and granulomatous amoebic encephalitis (GAE). In both diseases, treatment options are limited, and drug development is challenging. This study aimed to investigate the role of the large thioredoxin reductase selenoprotein of Acanthamoeba (AcTrxR-L) as a potential drug target assessing the effects of the thioredoxin reductase inhibitors auranofin, TRi-1, and TRi-2 on AcTrxR-L activity and on the viability of Acanthamoeba trophozoites. Recombinant expression and purification of AcTrxR-L as a selenoprotein allowed assessments of its enzymatic activity, with reduction of various substrates, including different thioredoxin isoforms. Auranofin demonstrated potent inhibition towards AcTrxR-L, followed by TRi-1, and TRi-2 exhibiting lower effectiveness. The inhibitors showed variable activity against trophozoites in culture, with TRi-1 and TRi-2 resulting in strongly impaired trophozoite viability. Cytotoxicity tests with human corneal epithelial cells revealed lower susceptibility to all compounds compared to Acanthamoeba trophozoites, underscoring their potential as future amoebicidal agents. Altogether, this study highlights the druggability of AcTrxR-L and suggests it to be a promising drug target for the treatment of Acanthamoeba infections. Further research is warranted to elucidate the role of AcTrxR-L in Acanthamoeba pathogenesis and to develop effective therapeutic strategies targeting this redox enzyme.

A Comprehensive Review on Acanthamoeba Keratitis: An Overview of Epidemiology, Risk Factors, and Therapeutic Strategies.

Acanthamoeba keratitis (AK) is a rare but severe corneal infection caused by the free-living amoeba, Acanthamoeba, which is ubiquitously present in the environment. This condition predominantly affects contact lens wearers but can also occur in non-lens users, particularly those exposed to contaminated water or with compromised immune systems. AK is characterized by progressive corneal inflammation, epithelial defects, and ulceration, which can lead to significant visual impairment or blindness if not promptly diagnosed and treated. This review aims to provide a comprehensive overview of AK by synthesizing current knowledge on its epidemiology, risk factors, pathophysiology, clinical manifestations, diagnostic approaches, and therapeutic strategies. The review also highlights preventive measures and public health strategies to reduce the incidence of this debilitating condition. A detailed examination of existing literature was conducted, focusing on the global incidence of AK, demographic trends, and various risk factors such as contact lens use, environmental exposures, and immunity status. The review also delves into the pathophysiology of Acanthamoeba infection, the host immune response, and the challenges in distinguishing AK from other forms of infectious keratitis. Therapeutic strategies, including medical and surgical interventions, are analyzed, along with emerging treatments. The global incidence of AK has increased, particularly among contact lens users, due to poor hygiene practices and environmental exposures. Early diagnosis remains challenging, often leading to delayed treatment and poorer outcomes. Biguanides and diamidines are the mainstays of medical therapy, with surgical options considered in advanced cases. Emerging therapies, such as photodynamic therapy and antimicrobial peptides, show promise in enhancing treatment outcomes.AK poses a significant threat to ocular health due to its potential for severe visual impairment and the complexities associated with its diagnosis and treatment. Early recognition, appropriate management, and public health initiatives focused on prevention are crucial for improving patient outcomes. Ongoing research and a collaborative approach among healthcare providers are essential to advancing the understanding and management of AK.

Assessing PCR-Positive Acanthamoeba Keratitis-A Retrospective Chart Review.

Ophthalmologists' diagnostic and treatment competence in Acanthamoeba keratitis varies widely. This investigator-initiated, retrospective, single-center chart review examined the electronic patient files regarding PCR-positive Acanthamoeba keratitis. We included corneal and contact lens assessments. We further reviewed the patient's medical history, corneal scraping results regarding viral or fungal co-infections, and the duration from symptom onset to final diagnosis. We identified 59 eyes of 52 patients from February 2010 to February 2023, with 31 of 52 (59.6%) being female patients. The median (IQR, range) patient age was 33 (25.3 to 45.5 [13 to 90]) years, and the mean (SD, range) time to diagnosis after symptom onset was 18 (10.5 to 35 [3 to 70]) days. Overall, 7 of 52 (7.7%) patients displayed a bilateral Acanthamoeba infection, and 48 (92.3%) used contact lenses at symptom onset. Regarding other microbiological co-infections, we found virologic PCR testing in 45 of 52 (86.5%) patients, with 3 (6.7%) positive corneal scrapings. Fungal cultures were performed in 49 of 52 (94.2%) patients, with 5 (10.2%) positive corneal scrapings. The medical treatment success rate was 45/46 (97.8%). This study raises awareness of patient education in contact lens handling and screens for further microbial co-infections in suspected Acanthamoeba cases.

Development of an Ex Vivo Porcine Eye Model for Exploring the Pathogenicity of Acanthamoeba.

Acanthamoeba, a widely distributed free-living amoeba found in various environments, is an opportunistic pathogen responsible for causing Acanthamoeba keratitis, a condition that may lead to blindness. However, identifying the pathogenicity of Acanthamoeba is challenging due to its complex life cycle, ability to adapt to different environments, variable virulence factors, and intricate interactions with the host immune system. Additionally, the development of an effective model for studying Acanthamoeba pathogenicity is limited, hindering a comprehensive understanding of the mechanisms underlying its virulence and host interactions. The aim of this study was to develop an ex vivo model for Acanthamoeba infection using porcine eyeballs and to evaluate the pathogenicity of the Acanthamoeba isolates. Based on slit lamp and biopsy analysis, the developed ex vivo model is capable of successfully infecting Acanthamoeba within 3 days. Histopathological staining revealed that clinical isolates of Acanthamoeba exhibited greater corneal stroma destruction and invasion in this model than environmental isolates. Our results highlight the importance of an ex vivo porcine eye model in elucidating the pathogenesis of Acanthamoeba infection and its potential implications for understanding and managing Acanthamoeba-related ocular diseases.

Novel anti-Acanthamoeba effects elicited by a repurposed poly (ADP-ribose) polymerase inhibitor AZ9482.

Acanthamoeba infection is a serious public health concern, necessitating the development of effective and safe anti-Acanthamoeba chemotherapies. Poly (ADP-ribose) polymerases (PARPs) govern a colossal amount of biological processes, such as DNA damage repair, protein degradation and apoptosis. Multiple PARP-targeted compounds have been approved for cancer treatment. However, repurposing of PARP inhibitors to treat Acanthamoeba is poorly understood. In the present study, we attempted to fill these knowledge gaps by performing anti-Acanthamoeba efficacy assays, cell biology experiments, bioinformatics, and transcriptomic analyses. Using a homology model of Acanthamoeba poly (ADP-ribose) polymerases (PARPs), molecular docking of approved drugs revealed three potential inhibitory compounds: olaparib, venadaparib and AZ9482. In particular, venadaparib exhibited superior docking scores (-13.71) and favorable predicted binding free energy (-89.28 kcal/mol), followed by AZ9482, which showed a docking score of -13.20 and a binding free energy of -92.13 kcal/mol. Notably, the positively charged cyclopropylamine in venadaparib established a salt bridge (through E535) and a hydrogen bond (via N531) within the binding pocket. For comparison, AZ9482 was well stacked by the surrounding aromatic residues including H625, Y652, Y659 and Y670. In an assessment of trophozoites viability, AZ9482 exhibited a dose-and time-dependent anti-trophozoite effect by suppressing Acanthamoeba PARP activity, unlike olaparib and venadaparib. An Annexin V-fluorescein isothiocyanate/propidium iodide apoptosis assay revealed AZ9482 induced trophozoite necrotic cell death rather than apoptosis. Transcriptomics analyses conducted on Acanthamoeba trophozoites treated with AZ9482 demonstrated an atlas of differentially regulated proteins and genes, and found that AZ9482 rapidly upregulates a multitude of DNA damage repair pathways in trophozoites, and intriguingly downregulates several virulent genes. Analyzing gene expression related to DNA damage repair pathway and the rate of apurinic/apyrimidinic (AP) sites indicated DNA damage efficacy and repair modulation in Acanthamoeba trophozoites following AZ9482 treatment. Collectively, these findings highlight AZ9482, as a structurally unique PARP inhibitor, provides a promising prototype for advancing anti-Acanthamoeba drug research.

Human Conjunctival Transcriptome in Acanthamoeba Keratitis: An Exploratory Study.

The purpose of this study was to identify conjunctival transcriptome differences in patients with Acanthamoeba keratitis compared with keratitis with no known associated pathogen. The host conjunctival transcriptome of 9 patients with Acanthamoeba keratitis (AK) is compared with the host conjunctival transcriptome of 13 patients with pathogen-free keratitis. Culture and/or confocal confirmed Acanthamoeba in 8 of 9 participants with AK who underwent metagenomic RNA sequencing as the likely pathogen. Cultures were negative in all 13 cases where metagenomic RNA sequencing did not identify a pathogen. Transcriptome analysis identified 36 genes differently expressed between patients with AK and patients with presumed sterile, or pathogen-free, keratitis. Gene enrichment analysis revealed that some of these genes participate in several biologic pathways important for cellular signaling, ion transport and homeostasis, glucose transport, and mitochondrial metabolism. Notable relatively differentially expressed genes with potential relevance to Acanthamoeba infection included CPS1 , SLC35B4 , STEAP2 , ATP2B2 , NMNAT3 , and AKAP12 . This research suggests that the local transcriptome in Acanthamoeba keratitis may be sufficiently robust to be detected in the conjunctiva and that corneas infected with Acanthamoeba may be distinguished from the inflamed cornea where no pathogen was identified. Given the low sensitivity for corneal cultures, identification of differentially expressed genes may serve as a suggestive transcriptional signature allowing for a complementary diagnostic technique to identify this blinding parasite. Knowledge of differentially expressed genes may also direct investigation of disease pathophysiology and suggest novel pathways for therapeutic targets.

Acanthamoeba Infection and Nasal Rinsing, United States, 1994-2022.

We describe 10 patients with nonkeratitis Acanthamoeba infection who reported performing nasal rinsing before becoming ill. All were immunocompromised, 7 had chronic sinusitis, and many used tap water for nasal rinsing. Immunocompromised persons should be educated about safe nasal rinsing to prevent free-living ameba infections.

Differentiation of acanthamoeba keratitis from other non-acanthamoeba keratitis: Risk factors and clinical features.

Infectious Keratitis is one of the most common ocular emergencies seen by ophthalmologists. Our aim is to identify the risk factors and clinical features of Acanthamoeba Keratitis (AK). This retrospective chart review study was conducted at King Khaled Eye Specialist Hospital, Riyadh, Saudi Arabia, and included all the microbial keratitis cases, male and female patients of all ages. The main outcome is the differentiation between various microbial keratitis types. We included 134 consecutive eyes of 126 persons. We had 24 cases of acanthamoeba keratitis, 22 bacterial keratitis, 24 fungal keratitis, 32 herpetic keratitis, and 32 bacterial co-infection. Contact lens wear was found in 33 eyes (24.6%). Among acanthamoeba keratitis patients, 73% were ≤ 39 years of age, and 73% were females (P <0.001). Also, in AK cases, epithelial defect was found in all cases (100%), endothelial plaques were found in 18 eyes (69.2%), 12 cases had radial keratoneuritis (46.2%), and ring infiltrate was found in 53.8% of AK cases. We determined the factors that increase the risk of acanthamoeba infection and the clinical characteristics that help distinguish it from other types of microbial keratitis. Our findings suggest that younger females and patients who wear contact lenses are more likely to develop acanthamoeba keratitis. The occurrence of epitheliopathy, ring infiltrate, radial keratoneuritis, and endothelial plaques indicate the possibility of acanthamoeba infection. Promoting education on wearing contact lenses is essential to reduce the risk of acanthamoeba infection, as it is the most significant risk factor for this infection.

Molecular detection and characterization of Acanthamoeba infection in dogs and its association with keratitis in Korea.

Acanthamoeba infection is associated with keratitis in humans; however, its association with keratitis in dogs remains unclear. To investigate this possibility, we collected 171 conjunctival swab samples from dogs with eye-related diseases (65 with keratitis and 106 without keratitis) at Chungbuk National University Veterinary Teaching Hospital, Korea, from August 2021 to September 2022. Polymerase chain reaction identified 9 samples (5.3%) as Acanthamoeba positive; of these, 3 were from dogs with keratitis (4.6%) and 6 were from dogs without keratitis (5.7%). Our results indicated no significant association between Acanthamoeba infection and keratitis, season, sex, or age. All Acanthamoeba organisms found in this study had the genotype T4, according to 18S ribosomal RNA analysis. Acanthamoeba infection in dogs might have only a limited association with keratitis.

Could giant viruses be considered as a biotechnological tool for preventing and controlling Acanthamoeba infections?

The aim of the study was to evaluate the efficiency of mimivirus as a potential therapeutic and prophylactic tool against Acanthamoeba castellanii, the etiological agent of Acanthamoeba keratitis, a progressive corneal infection, that is commonly associated with the use of contact lenses and can lead to blindness if not properly treated. Mimivirus particles were tested in different multiplicity of infection, along with commercial multipurpose contact lenses' solutions, aiming to assess their ability to prevent encystment and excystment of A. castellanii. Solutions were evaluated for their amoebicidal potential and cytotoxicity in MDCK cells, as well as their effectiveness in preventing A. castellanii damage in Madin-Darby canine kidney (MDCK) cells. Results indicated that mimivirus was able to inhibit the formation of A. castellanii cysts, even in the presence of Neff encystment solution. Mimivirus also showed greater effectiveness in controlling A. castellanii excystment compared to commercial solutions. Additionally, mimivirus solution was more effective in preventing damage caused by A. castellanii, presented greater amoebicidal activity, and were less cytotoxic to MDCK cells than commercial MPS. Mimivirus demonstrates a greater ability to inhibit A. castellanii encystment and excystment compared to commercial multipurpose contact lens solutions. Additionally, mimivirus is less toxic to MDCK cells than those commercial solutions. New studies utilizing in vivo models will be crucial for confirming safety and efficacy parameters.

Lactase can target cellular differentiation of Acanthamoeba castellanii belonging to the T4 genotype.

The free living Acanthamoeba spp. are ubiquitous amoebae associated with potentially blinding disease known as Acanthamoeba keratitis (AK) and a fatal central nervous system infection granulomatous amoebic encephalitis (GAE). With the inherent ability of cellular differentiation, it can phenotypically transform to a dormant cyst form from an active trophozoite form. Acanthamoeba cysts are highly resistant to therapeutic agents as well as contact lens cleaning solutions. One way to tackle drug resistance against Acanthamoeba is by inhibiting the formation of cysts from trophozoites. The biochemical analysis showed that the major component of Acanthamoeba cyst wall is composed of carbohydrate moieties such as galactose and glucose. The disaccharide of galactose and glucose is lactose. In this study, we analyzed the potential of lactase enzyme to target carbohydrate moieties of cyst walls. Amoebicidal assessment showed that lactase was ineffective against trophozoite of A. castellanii but enhanced amoebicidal effects of chlorhexidine. The lactase enzyme did not show any toxicity against normal human keratinocyte cells (HaCaT) at the tested range. Hence, lactase can be used for further assessment for development of potential therapeutic agents in the management of Acanthamoeba infection as well as formulation of effective contact lens disinfectants.

Microbial keratitis in Sao Paulo, Brazil: a 10-year review of laboratory results, epidemiological features, and risk factors.

To study epidemiological data, laboratory results, and risk factors associated with microbial keratitis. We conducted a retrospective study of corneal sample cultures from patients with microbial keratitis from January 2010 to December 2019. Results were analyzed according to the etiological diagnosis of bacterial, mycotic, or parasitic infection and were associated with related risk factors. We analyzed 4810 corneal samples from 4047 patients (mean age 47.79 ± 20.68 years; male 53.27%). The prevalence of bacterial, fungal, and Acanthamoeba infections were 69.80%, 7.31%, and 3.51%, respectively. The most frequently isolated bacteria were coagulase-negative Staphylococcus (CoNS) (45.14%), S. aureus (10.02%), Pseudomonas spp. (8.80%), and Corynebacterium spp. (6.21%). Among CoNS, the main agent was S. epidermidis (n=665). For mycotic keratitis, Fusarium spp. (35.42%) and Candida parapsilosis (16.07%) were the most common agents among filamentous and yeasts isolates, respectively. Contact lens use was associated with a positive culture for Acanthamoeba spp. (OR = 19.04; p < 0.001) and Pseudomonas spp. (OR = 3.20; p < 0.001). Previous ocular trauma was associated with positive fungal cultures (OR = 1.80; p = 0.007), while older age was associated with positive bacterial culture (OR = 1.76; p = 0.001). Our findings demonstrated a higher positivity of corneal sample cultures for bacteria. Among those, CoNS was the most frequently identified, with S. epidermidis as the main agent. In fungal keratitis, Fusarium spp. was the most commonly isolated. Contact lens wearers had higher risks of positive cultures for Acanthamoeba spp. and Pseudomonas spp. Ocular trauma increased the risk of fungal infection, while older age increased the risk of bacterial infection.